KATHRYN MEDLER LAB

Why should we care about taste?
Taste is used to determine if potential food items will be ingested or rejected.
Taste is used by all organisms and is the oldest sensory system.
Loss of taste can lead to depression and loss of appetite which can cause malnutrition. Deficits in the taste system can also lead to uncontrolled appetite and obesity.

Mammalian taste buds are present in papillae on the tongue
Apical
Taste stimuli
Taste bud
Lingual
Epithelium
Basolateral
Afferent gustatory nerve fiber

Taste Transduction: Two distinct signaling pathways exist
Na + H +
+
+
+
Bitter
Sweet
Umami
R
PLC
IP
3
Multiple signaling pathways are present taste cells and all pathways depend on increases in intracellular calcium to transmit signals to the nervous system.
Ca 2+
Store
Na +
K +
[Ca 2+ ] i
Serotonin
Ca 2+
[Ca 2+ ] i
ATP
TRPM5
Our lab is studying these different signaling mechanisms: how they function and how they are regulated.

Calcium imaging measures changes in calcium levels in live cells
We can characterize the functional role of different proteins expressed in these cells and how they affect calcium signals.

1750
1500
1250
1000
750
500
250
0
0
“Complex” stimuli
Bitter hi K
100
Time (s)
200 300
“Ionic” stimuli
1750 Bitter hi K
1500
1250
1000
750
500
250
0
0 100 200
Time (s)
300 400
Activate calcium release from stores Activate voltage gated calcium influx
Different taste stimuli evoke different signals.
Hacker et al., 2008

1250
Bitter hi K
1000
750
500
250
0
0 200
Time (s)
400
Some taste cells responded to bitter stimuli AND cell depolarization.
600
Hacker et al., 2008
This is a newly identified sub-population of taste cells.

How do these taste cells respond to multiple stimuli?

Expression patterns of PLC b
3/IP
3
R1 in taste cells
D
PLC b
3 and IP
3
R1 are co-expressed in a population of taste cells that are distinct from the PLC b
2 expressing cells.
Further studies are being conducted to characterize this newly identified signaling pathway.

There are 3 separate taste cell groups.
+
+
+ +
+ +
Na +
Ca 2+

[Ca 2+ ] i i
 gustducin
Phospholipase C b
2
+
+
+
+
+
Na +
G proteins
Phospholipase C b
3
IP
3
R1
IP
3
Ca 2+
Endoplasmic reticulum
 [Ca 2+ ] ] i i i
Ca 2+
We are asking “How do each of these groups contribute to detection of taste stimuli?”

We are also studying the evoked taste responses in obese mice.
We asked “Are peripheral taste responses different in obese mice versus normal mice?”
Norm
Obese 75
100
Norm
Obese
NS
75
50
NS
50
25
***
***
25 ***
**
*** ***
0
MPG Sac Ace K Den hi K
0
MPG Sac AceK Den
The number of responsive taste cells and the response amplitudes are reduced in obese mice for the appetitive tastes.
We’re asking how does this affect the animal’s ability to perceive taste stimuli? Is it reversible?

TRPM5 is a well-known monovalent selective TRP channel that is important in taste transduction. TRPM5 turns on in response to some taste stimulation.
We found that taste cells also express TRPM4, which is the other monovalent selective TRP channel. TRPM4 is also activated by some taste stimuli.

110
DEN
Ca
2+
Na
+
20
100
90
0 100
Hi K
200
Time (s)
300 400
10
0
In some cells, taste stimuli evoke sodium and calcium increases. Using imaging and patch clamp, we’re determining how
TRPM4 contributes to these responses.

Gene regulation by WT1 in taste cells
In collaboration with Stefan Roberts lab

Transcriptional Regulation by WT1
BASP1
WT1
General transcription machinery
IIA
IIF
IIH
IIB
IIE
Pol II
IID
TATA
Growth factors
Amphiregulin
IGFII
PDGF-A
Apoptosis
Bcl 2
Bak c-myc
Differentiation
Podocalyxin
Nephrin

WT1 plays a critical role in the development of several organs and tissues
WT1 Knock-out mice
Kidneys
Gonads
Spleen
Adrenal glands
Diaphragm
Retinal Ganglia
Olfactory epithelium
Taste buds

WT1 and BASP1 are expressed in taste cells
Adult Embryonic
WT1
WT1 Ctrl
BASP1 Ctrl
E13 E14.5
E17.5

WT1 null mice fail to develop a peripheral taste system
E13.5
WT WT
Troma 1 Sox2
KO
WT
KO
WT
GAP-43 Shh
KO KO

WT1 regulates genes critical for taste cell development
Real time PCR
CHiP assay
1,2
1
0,8
0,6
0,4
0,2
0
* LEF1
4
3
2
1
0
6
5
LEF1
IgG WT1 BASP1
WT
1,2
1
0,8
0,6
0,4
0,2
0
WT
KO
*
PTCH1
7
6
5
4
3
2
1
0
PTCH1
IgG WT1 BASP1
KO
1,2
1
0,8
0,6
0,4
0,2
0
*
WT1+/+ WT1-/-
BMP4

Primary taste cells can be cultured and transfected
Hoechst
WT1
Hoechst
PLC β2
Hoechst
TRMP5-GFP
3,5
3
2,5
2
1,5
1
0,5
0
14
12
10
8
6
4
2
0
LEF1
IgG
PTCH1
IgG
CHiP
WT1
WT1
BASP1
BASP1
Knockdown of WT1 in cultured taste buds causes a reduction in the expression of WT1 target genes that are important in taste cell maintenance.
qPCR
1,2
1
0,8
0,6
0,4
0,2
0
1,2
1
0,8
0,6
0,4
0,2
0
1,2
1
0,8
0,6
0,4
0,2
0
WT1
Control siRNA
WT1 siRNA
LEF1
Control siRNA
WT1 siRNA
PTCH1
Control siRNA
WT1 siRNA

Combine the physiological and molecular approaches of the Medler and Roberts labs to study the role of
WT1 and BASP1 in gene regulation during development and tissue homeostasis

If you are interested in rotating in the lab on any of these projects, please contact me by email: kmedler@buffalo.edu